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Sebaihia M, Peck MW, Minton NP, et al. Genome sequence of a proteolytic (Group I) Clostridium botulinum strain Hall A and comparative analysis of the clostridial genomes. Genome Research. 2007;17(7):1082-1092. doi:10.1101/gr.6282807.

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Grow in soil

Clostridium botulinum spores are ubiquitous in the environment. They are found in the soil and have been recovered in agricultural products such as honey. Spores are also found in marine sediments and the intestinal tracts of animals. The spores are highly resistant to inactivation. As a result, control methods for botulism focus on inhibiting the organism's growth and toxin production, rather than treating the food product to eliminate the spores.

Grow in canned food pH less than 4.5

Home-canned products, especially of low-acid foods, are attributed to most cases of foodborne botulism, because the time and temperature to which the food is heated are often inadequate. Susceptible foods include canned asparagus, green beans, garlic in oil, corn, soups, ripe olives, tuna, sausage, luncheon meats, fermented meats, salad dressings, and smoked fish. Additionally, spores of C. botulinum have been found on the surfaces of vegetables and fruits, and infant botulism has been linked to the ingestion of spores in honey, corn syrup, and other foods (FDA 2012). B. coagulans was inoculated into sterile, tubed tomato juice and growth and pH were observed for 21 days. Juice samples initially at pH 4.5 supported good growth; pH was elevated to 5.0 after 6 days incubation at 35°C. Other tomato juice samples (initial pH 5.0 or 5.5) also supported good growth of these flat-sour bacteria and with corresponding increases in pH. The results indicate that flat-sour spoilage of tomatoes and similar foods may cause decreased acidity, thus possibly allowing outgrowth of Clostridium spores in these water bath-processed foods, thus making  it is possible for C. botulinum to grow.

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Taxonomy:

Domain  : Bacteria

Phylum   : Firmicutes

Class      : Clostridia

Order     : Clostridiales

Family    : Clostridiaceae

Genus    : Clostridium

Species  : C. botulinum

Characteristic and morphology:
Clostridium botulinum is a gram positive, rod shaped foodborne pathogen that can form spore to keep survive in harsh environment. During sporulation process,  this bacteria will produce botulinum neurotoxin on the food that can cause Botulism disease to human or animal that consume the contaminated food.

C. botulinum is motile due to the presence of flagella and cannot tolerate to the presence of oxygen. This obligate anaerobic bacteria cannot survive in oxygen due to the absence of enzyme superoxide dismutase on their cell. This enzyme is important as it can convert the lethal superoxide that formed in bacteria cell in the presence of oxygen.

This bacteria can be classified into 4 groups:
- Group 1 (proteolytic) and Group 2 (non-proteolytic) can cause Boltulism disease in human that consume contaminated food.
- Group 3 is the group of Clostridium botulinum that can cause disease to animal.
-Group 4 is not the type of this bacteria that can cause disease to human or animal.  There are no research yet that can proof this group can cause harm to human or animal body.

This bacteria can be isolated in haemolysis blood agar and egg yolk agar. Morphology of the colony that form in these agar can help the microbiologist to differentiate the group of Clostridium botulinum bacteria.

Colony morphology in blood agar
Group 1

2—6 mm diameter

irregular margin,

opaque, raised center

narrow hemolysis

Group 2

1-3 mm diameter

slightly irregular with lobate margins

translucent to semiopaque with matt surface

mosaic structure in oblique light

narrow hemolysis

Colony morphology in egg yolk agar
Group 1 and 2

Iridescent layer on and around colonies

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ECOLOGY (butolinum type that cause disease)

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ECOLOGY (butolinum type that cause disease)

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CAN BE CLASSIFIED INTO:
1)TYPE A STRAIN.
Type A is one of the most common serotypes that cause human botulism in several countries and numerous investigations have focused on the genetic diversity of this serotype (Luquez et. Al, 2012). This type are inhibited by soil isolates of C. sporogenes, C. perfringens and B. cereus.
2) TYPE B STRAIN
This types produces neurotoxin which is initially of low toxicity but which then becomes activated to highly toxic form by the action of an endogenous enzyme(s) (Dasgupta,1971). The toxins produced by these strains, however, are regarded as physicochemically and immunologically identical.  Clostridium botulinum type B involves proteolytic and nonproteolytic strains, being different from each other also in other biological properties.
3) TYPE E STRAIN
inhibited by other nontoxic organisms whose biochemical properties and morphological characteristic were similaR to type E. The sources for this strain is from marime mamals.
4) TYPE F STRAIN
in recent studies, this type absence in mud sample due to the presence of Bacillus licheniformis because this bacillus inhibit type F strains.

REFERENCE
1) Lúquez, C., Raphael, B. H., Joseph, L. A., Meno, S. R., Fernández, R. A., & Maslanka, S. E. (2012). Genetic Diversity among Clostridium botulinum Strains Harboring bont/A2 and bont/A3 Genes. Applied and Environmental Microbiology, 78(24), 8712–8718. http://doi.org/10.1128/AEM.02428-12
2)DasGupta, B. R. (1971). Activation of Clostridium botulinum Type B Toxin by an Endogenous Enzyme. Journal of Bacteriology, 108(3), 1051–1057.

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Signs and symptoms of foodborne botulism typically begin between 12 and 36 hours after the toxin gets into your body. But, the start of symptoms can range from a few hours to several days, depending on the amount of toxin ingested
-difficulty in swallowing and speaking
-naussea
-vomitting
-diarrhea
-double vision or blurred
-dry mouth
-paralysis
Clostridium botulinum also can cause infant botulism.
If infant botulism is related to food, such as honey, problems generally begin within 18 to 36 hours after the toxin enters the baby's body. Signs and symptoms include:

Constipation (often the first sign)

Floppy movements due to muscle weakness and trouble controlling the head

Weak cry

Irritability

Drooling

Drooping eyelids

Tiredness

Difficulty sucking or feeding

Paralysis

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1. CNN- Federal Bureau of Prisons believed that the botulism is the cause of severe illness that affected 20 inmates at a federal prison at Mississippi.

They became ill as a result of drinking homemade alcohol

an inmate ill as a result of drinking tainted moonshine, (Oklahoma City)

#who consumed the contraband beverages are at risk.

2. Farfield Medical center (FMC) and Farfield Department of Health contacted Ohio Department of Health (ODH)-

All affected person had eaten on the church potluck meal. Based on the interview from the person that ate at the church potluck and said that the consumption of potato salad (home made) is the highest probability that the affected person ate. The attendee who prepared the potato salad said they just use a boiling water canner which does not kill the botulinam spores. The potatoes also does not being heated after the removal of the can.

3. Cincinnati Health Department was notified-

Patient A had been treated for 12 days for suspected autoimmune disease. When patient B, the roommate of patient A, was evaluated at the same medical center for similar symptoms. They have been investigated that both Patient A and B, shared a meal that included prepackaged pesto from a jar. After performing ELISA test, it was observed that there is the presence of the botulinum toxin type-B. Environmental assessment of company that they buy the jar identified improper acidification and pressurization practices and lack of licensure to sell canned products commercially.

4. All India Institute of Medical Sciences received serum and food samples to detect possible outbreak that can be happen in India-

34 of 310 students of a residential school in rural Gujrat complained of abdominal pain, nausea, chest pain, and difficulty in breathing.Patients reported that 24 hours before onset of symptoms, they had eaten ladoo (a local sweet), curd, buttermilk, sevu (crisp made of gram flour), and pickle.

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C. botulinum is found worldwide and cases of botulism can be seen anywhere. In ruminants, botulism mainly occurs in areas where phosphorus or protein deficiencies are found. Botulism is seen regularly in cattle in South Africa and sheep in Australia. This disease is rare in ruminants in the United States, although a few cases have been reported in Texas and Montana.

The species of Clostridia associated with animal infections have been isolated from a wide spectrum of environmental sources which dust, sewage, rivers, lakes, sea water, milk, vegetables, fresh meat, insects and fish, in addition to soils and the intestinal tract. Therefore, Clostridia of veterinary importance can be expected to be recovered from any clinical specimen contaminated by soil, dust or faeces. As members of the normal or transient flora of the respiratory and genito-urinary tracts, they ma y be expected to be recovered from anybody fluid or tissue contaminated colonized, or infected by these endogenous sources.

In the spore stage, all members of the genus Clostridium have a pronounced but variable resistance to heat, drying and disinfectants. Thus, they are able to survive in the environment under favorable conditions. In the vegetative state, clostridia are about as resistant to physical and chemical agents and unfavorable environmental conditions as non-sporing aerobes.

Although clostridia usually lead a saprophytic existence, several species are causally related to well-recognized diseases in animals and Man. Some clostridia produce powerful exotoxin which are responsible for the specific pathogenicity of the organisms. Others may produce less potent toxins but, in addition, have the power of invading and multiplying in th tissues, causing wound infections such as gas gangrene.

https://www.oie.int/doc/ged/D8192.PDF

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Most of food outbreak that was caused by Clostrodium botulinum are from the Group 1 which produce toxin types A, B and F. Production of extracellular proteases by proteolysis has been associated with food spoilage and pathogenicity. Besides that, it also metabolise chitin, lipase and partially metabolise starch.

Amino acids.

The proteinaceous products, peptides and amino acids that areproduced by the C. botulinum extracellular proteases. It cleave the surrounding polypeptides so the bacteria can digest it into smaller peptides.

It uses fermentation pathways that utilize a series of coupled oxidation-reduction reactions to acquire energy, which is the Strickland Reaction. The oxidation of one amino acid is directly coupled to the reduction of another amino acid. Some of the amino acids that are used as oxidised reagent are alanine, leusine, isoluecine, and valine. Other amino acids are used as reducing agent such as glycine, proline, hydroxyproline, and orthinine. The reaction results in the production of CO₂ and a nitrogen source, most commonly NH3.

The amino acid glycine is first reduced by a glycine reductase complex and then oxidized by a glycine cleavage system (Sebaihia et al. 2007).

Starch

Starch is another abundant polysaccharide that consists of a linear polymer, amylose, and a branched polymer, amylopectin. Complete degradation of starch requires the combined action of several enzymes, alpha-amylase, beta-amylase, pullulanase, and glucoamylase but C. botulinum only able to secrete beta-amylase.

This enzyme catalyzes the removal of maltose molecules from the nonreducing ends of the starch polymers. The lack of other starch-hydrolyzing enzymes in C. botulinum suggests that this bacterium is unable to completely degrade whole starch but nevertheless can degrade starch-derived polysaccharides.

Lipase

Two genes C. botulinum codes for two putatively secreted lipases. Lipase activity leads to the production of a thin pearly layer on and around colonies formed on media containing egg yolk.

It can be used as a simple diagnostic test for the detection of C. botulinum.

Chitin

Clostridium botulinum uses the second most abundant sugar, as its second main source of energy.

Clostridium botulinum’s genomeencodes five different enzymes that are capable of metabolizing this sugar. In addition to being a source of energy, chitin can also provide supplementary carbon and nitrogen (Peck et al. 2010).

Figure 1. Chitin degradation by Clostridium Botulium http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1899119/.

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